Role of the Serine-Rich Surface Glycoprotein Srr1 of Streptococcus agalactiae in the Pathogenesis of Infective Endocarditis
Sullam PM (2013) Role of the Serine-Rich Surface Glycoprotein Srr1 of Streptococcus agalactiae in the Pathogenesis of Infective
Endocarditis. PLoS ONE 8(5): e64204. doi:10.1371/journal.pone.0064204
Role of the Serine-Rich Surface Glycoprotein Srr1 of Streptococcus agalactiae in the Pathogenesis of Infective Endocarditis
Ho Seong Seo 0
Yan Q. Xiong 0
Paul M. Sullam 0
Robert A. Burne, University of Florida, United States of America
0 1 Division of Infectious Diseases, Veterans Affairs Medical Center and the University of California San Francisco , San Francisco , California, United States of America, 2 Department of Medicine, Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, California, United States of America, 3 Geffen School of Medicine at UCLA , Los Angeles, California , United States of America
The binding of bacteria to fibrinogen and platelets are important events in the pathogenesis of infective endocarditis. Srr1 is a serine-rich repeat glycoprotein of Streptococcus agalactiae that binds directly to the Aa chain of human fibrinogen. To assess the impact of Srr1 on the pathogenesis of endocarditis due to S. agalactiae, we first examined the binding of this organism to immobilized human platelets. Strains expressing Srr1 had significantly higher levels of binding to human platelets in vitro, as compared with isogenic Dsrr1 mutants. In addition, platelet binding was inhibited by pretreatment with anti-fibrinogen IgG or purified Srr1 binding region. To assess the contribution of Srr1 to pathogenicity, we compared the relative virulence of S. agalactiae NCTC 10/84 strain and its Dsrr1 mutant in a rat model of endocarditis, where animals were co-infected with the WT and the mutant strains at a 1:1 ratio. At 72 h post-infection, bacterial densities (CFU/g) of the WT strain within vegetations, kidneys, and spleens were significantly higher, as compared with the Dsrr1 mutant. These results indicate that Srr1 contributes to the pathogenesis of endocarditis due to S. agalactiae, at least in part through its role in fibrinogen-mediated platelet binding.
Funding: This study was supported by the Department of Veterans Affairs and the VA Merit Review program, the Northern California Institute for Research and
Education, NIH grants R01-AI41513 (P.M.S.), R01-AI057433 (P.M.S.), and a Fellowship Award from the American Heart Association, Western Affiliate (H.S.S). The
funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
Streptococcus agalactiae (Group B streptococcus [GBS]) is a
frequent cause of neonatal meningitis and sepsis. In recent years,
however, GBS infections in nonpregnant adults are being
increasingly reported. Individuals at greater risk for this disease
include the elderly, immunosuppressed patients, and diabetics [1
3]. Although GBS is a relatively uncommon cause of endocarditis
(accounting for 12% of culture-positive cases), endovascular
infection due to this organism is associated with a high mortality
rate (3450%), especially in the setting of prosthetic valve infection
. Complications such as sepsis, valvular destruction, cardiac
failure, and embolic phenomena are also frequent in this disease
The pathogenesis of endocarditis is a complex process, involving
multiple host-pathogen interactions. A central aspect of virulence
in this disease is the ability of organisms to bind host components,
such as fibrinogen, fibronectin, and platelets . These
binding events appear to be important both for the initial
attachment of bacteria to the endovascular surface, and for the
subsequent progression of infection. For several Gram-positive
bacteria, binding to human platelets is mediated in part by an
adhesin belonging to the serine-rich repeat (SRR) glycoprotein
. For example, strains of Streptococcus gordonii can bind
platelets directly via the interaction of the SRR adhesins GspB or
Hsa with a receptor (GPIb) on the platelet membrane . The
binding of Staphylococcus aureus to platelets is mediated in part by the
SRR protein SraP, though the receptor for this adhesin remains
unidentified . In addition, S. aureus can attach to platelets via
fibrinogen and fibrin, which act as a molecular bridge between the
bacteria and platelet surface .
Two SRR glycoproteins (Srr1 and Srr2) have been identified
thus far in S. agalactiae . Expression of Srr1 has been shown to
contribute to colonization and virulence in models of GBS
bacteremia and meningitis infection . In addition, we have
recently demonstrated that Srr1 binds to human fibrinogen via its
interaction with the Aa chain of the protein, and that loss of
fibrinogen binding is associated with decreased attachment to
brain microvascular endothelial cells in vitro, as well as attenuated
virulence, in an experimental model of meningitis . In view of
the importance of fibrinogen binding for endovascular infection,
we examined the impact of Srr1 on platelet binding in vitro, and its
role in the pathogenesis of infective endocarditis.
Materials and Methods
Purified human fibrinogen was obtained from Haematologic
Technologies. Rat fibrinogen was purchased from Sigma-Aldrich.
was acquired from Innovative
Strains, plasmids, and growth conditions
The bacteria and plasmids used in this study are listed in
Tables 1. S. agalactiae strains were grown in Todd-Hewitt broth
(Difco) supplemented with 0.5% yeast extract (THY broth). All
mutant strains grew at a comparable rate in vitro as compared with
respective parental strains (data not shown). Escherichia coli strains
DH5a, BL21 and BL21(DE3) were grown at 37uC under aeration
in Luria Bertani broth (LB; Difco). Antibiotics were added to the
media as required. All isolates were stored at 280uC until thawed
just prior to use.
Cloning and expression of the Srr1 binding region
Genomic DNA was isolated from GBS NCTC 10/84 using
Wizard Genomic DNA purification kits (Promega), according to
the manufacturers instructions. PCR products were cloned into
either pET28FLAG or pET22(+) to express FLAG-tagged or
His6tagged versions of Srr1-BR (amino acids [AA] 303641 of the
SRR1). Proteins were purified by either Ni-NTA (Promega) or
anti-FLAG M2 agarose affinity chromatography (Sigma-Aldrich).
Binding of S. agalactiae to immobilized human platelets
and rat fibrinogen
Overnight cultures of S. agalactiae were harvested by
centrifugation, washed in PBS, and adjusted to a concentration of
106 CFU/ml. Purified rat fibrinogen or washed, fixed human
platelets were immobilized in 96-well microtiter plates as described
previously (46). The plates were then treated with casein-based
blocking solution (Roche) at 37uC for 1 h and washed three times
with PBS. Purified recombinant Srr1-BR (200 mg/ml), anti-Srr1
IgG (100 mg/ml), or anti-fibrinogen IgG (100 mg/ml) were added
to the wells for 30 min, followed by washing and the addition of
100 ml of the bacterial suspension. The plates were incubated at
room temperature for 1 h, and the wells were washed three times
with PBS to remove nonadherent organisms. The wells were
treated with 100 ml of trypsin (2.5 mg/ml) for 10 min at 37uC to
release the attached bacteria, and the number of bound bacteria
was determined by plating serial dilutions of the recovered bacteria
onto blood agar plates as previously described .
Binding of recombinant Srr1-BR to human platelets
Fixed human platelets were immobilized in 96 well cell culture
plates as described previously . After treatment with a
caseinbased blocking reagent, the wells were incubated with
FLAGSrr1BR (04 mM) in PBS for 1 h at room temperature, followed by
washing. Bound protein was detected by ELISA with anti-FLAG
monoclonal antibody. For some studies, the wells were
preincubated with His6-tagged Srr1-BR (050 mM) or anti-fibrinogen IgG
(0100 mg/ml) for 0.5 h at RT followed by washing, prior to
adding FLAGSrr1-BR (1 mM).
Rat model of infective endocarditis
The relative virulence of S. agalactiae NCTC 10/84 parental
strain and its isogenic variant (NCTC 10/84 Dsrr1) was assessed in
a competition model of IE in rats, as described previously [29,30].
In brief, Sprague-Dawley female rats (250 to 300 g, Harlan
Laboratory, Inc.) were first anesthetized with ketamine (35 mg/kg)
and xylazine (10 mg/kg). A sterile polyethylene catheter was
surgically placed across the aortic valve of each animal, such that
the tip was positioned in the left ventricle, to induce the formation
of sterile valvular vegetations (nonbacterial thrombotic
endocarditis) [27,29]. The catheters were left in place throughout the
study. Three days post-catheterization, the animals were infected
intravenously with an inoculum of approximately 56105 CFU
containing a 1:1 mixture of S. agalactiae NCTC 10/84 and its Dsrr1
isogenic variant. At 72 hr post-infection, the rats were euthanized
with thiopental (100 mg IP). Animals were included in the final
analysis only if the catheters were correctly positioned across the
aortic valve at the time of sacrifice, and if macroscopic vegetations
were visible. At sacrifice, all cardiac vegetations, as well as kidneys
and spleens, were harvested, weighed, homogenized in saline,
Genotype or descriptiona
expression host, inducible T7 RNA polymerase
serotype III, clinical isolate
serotype V, clinical isolate
NCTC 10/84Dsrr1, CmR
streptococcal shuttle vector, ErmR
vector for expression of Srr1, ErmR
expression vector, AmpR
expression vector with FLAG-tag, KanR
vector for expression of Srr1-BR, AmpR
vector for expression of FLAG-tagged Srr1303641
aCmR, chloramphenicol resistance; ErmR, erythromycin resistance; AmpR, ampicillin resistance; KanR, kanamycin resistance.
Figure 1. GBS binding to immobilized human platelets is mediated by glycoprotein Srr1. (A) Platelet binding by GBS strains COH31 and
NCTC 10/84, their Dsrr1 isogenic variants, and the mutant strains complemented in trans with srr1 (pSrr1). (B) GBS binding to human platelets was
inhibited by pretreating the monolayers with 100 mg/ml of anti-fibrinogen IgG (Anti-Fg). Normal IgG (IgG) served as a control. (C) Inhibition of
binding by recombinant Srr1 binding region (Srr1-BR). Levels of binding were calculated as relative to the WT strains (mean 6 SD). Values shown
represent the means (6 S.D.) of triplicate measurements. * = P,0.01.
serially diluted, and plated onto 8% sleep blood Todd Hewitt agar
(with or without 2.5 mg/ml of chloramphenicol) for quantitative
culture. The plates were incubated for 48 h at 37uC, and bacterial
densities were expressed as the log10CFU per gram of tissue.
Differences in means +/2 SD were compared for statistical
significance by the paired t-test. The data were also analyzed by
calculating a competition index, which was defined as the ratio
of the paired strains within tissues for each animal, normalized by
the ratio of organisms in the inoculum [27,29]. The mean of the
log10 normalized ratios was tested against the hypothesized no
effect mean value of 0, using a paired t-test, with P#0.05 as the
threshold for statistical significance.
Animals were maintained in accordance with the American
Association for Accreditation of Laboratory Animal Care
(AAALAC) criteria. All animal studies were approved by the
Animal Research Committee (IACUC) of the Los Angeles
Biomedical Research Institute at Harbor-UCLA Medical Center.
Binding of GBS to human platelets is mediated by
To assess whether GBS binding to human platelets is mediated
by Srr1, we compared two GBS strains (COH31 and NCTC 10/
84) and their respective srr1 deletion variants for adherence to
these cells in vitro (Fig. 1A). Both strains bound platelets
significantly above background levels, with 28.062.6% and
12.062.6% (mean 6 SD) of the inoculum bound for COH31
and NCTC 10/84, respectively. Levels of binding by both srr1
mutant strains were significantly lower than those of the parent
strains, with a 79.263.4% reduction in platelet binding for
COH31Dsrr1 and a 71.464.2% reduction for NCTC 10/84Dsrr1.
Figure 2. Recombinant Srr1-BR interacts with human platelets. (A) Binding of FLAGSrr1-BR protein to immobilized platelets. (B) Inhibition of
FLAGSrr1-BR binding to platelets by His6 tagged Srr1-BR. Platelets were pretreated with the indicated concentrations of His6 tagged Srr1-BR. (C)
Binding of FLAGSrr1-BR to immobilized platelets pretreated with anti-fibrinogen IgG or preimmune rabbit IgG. Values represent relative binding of
FLAGSrr1-BR binding as compared with untreated platelets. Bars indicate the means (6 S.D.). * = P,0.01.
Complementation of the srr1 mutation in trans restored binding by
both mutant strains, thereby demonstrating that the loss of binding
observed with srr1 disruption was not due to polar or pleiotropic
effects. In addition, GBS binding to human platelets was inhibited
by rabbit anti-fibrinogen IgG, but not by normal rabbit IgG
(Fig. 1B), with WT GBS binding levels reduced to those seen with
the srr1 deletion strains. We then examined the impact of
preincubating the platelet monolayers with the recombinant
binding domain of Srr1 (Srr1-BR). As shown in Fig. 1C,
pretreating the immobilized platelets with recombinant Srr1-BR
inhibited subsequent binding by both GBS strains. Since previous
studies have shown that human platelets express
membraneassociated fibrinogen [11,31,32], our results indicate that GBS
binding to human platelets is mediated by the interaction of Srr1
with fibrinogen on the surface of these cells.
Binding of Srr1-BR to immobilized platelets
To further assess the role of Srr1, we evaluated the binding of
FLAG-tagged Srr1-BR (FLAGSrr1-BR) to immobilized human
platelets. We found that FLAGSrr1-BR interacted with platelets in a
concentration-dependent manner, when tested over a range of 0
4 mM (Fig. 2A). In addition, binding was significantly inhibited by
preincubating the platelets with His6-tagged Srr1-BR, (Fig. 2B) or
anti-fibrinogen IgG (Fig. 2C). These results demonstrate that
Srr1BR can bind platelets via its interaction with fibrinogen, and that
this interaction is specific.
Effect of Srr1 expression on streptococcal endocarditis
Some fibrinogen binding proteins, such as ClfA of S. aureus, bind
fibrinogen from only certain animal species . With a view
towards in vivo studies, we next sought to assess whether Srr1 had a
similar impact on the interaction of GBS with rat fibrinogen.
PSIBLAST analysis indicated that the predicted binding region in rat
fibrinogen is located at AA294334 of the Aa chain, which has
49% identity with the Srr1 binding site on human fibrinogen
(Fig. 3A). When tested in vitro, binding of the isogenic mutants to
rat fibrinogen was found to be significantly lower than that of wild
type strains COH31 and NCTC 10/84 (Fig. 3B). In addition,
FLAGSrr1-BR was bound to immobilized rat fibrinogen in a
concentration-dependent manner, as was seen previously with
human fibrinogen (Fig. 3C) .
To examine the impact of Srr1 expression on the pathogenesis
of endocarditis, we compared the relative virulence of GBS NCTC
10/84 with its isogenic mutant (Dsrr1), as measured by a rat
coinfection model of this disease. Animals (n = 14) had significantly
lower densities of the mutant strain (mean log10 CFU/g 6
SD = 7.4661.63) within vegetations as compared with the parent
strain (8.6261.25). Levels of the mutant strain were also
significantly reduced within kidneys and spleens (Table 2). We
then re-analyzed these data by comparing the ratio of the isogenic
strains within tissues, with the CFU of each strain normalized to
the number of CFU within the inoculum (competition index)
(Fig. 4). When assessed by this approach, the levels of the srr1
mutant (Dsrr1) remained significantly reduced in all tissues, as
compared with WT. Thus, Srr1 appears to be a significant
virulence determinant for the pathogenesis of endocarditis due to
Mean S.D. (log10CFU/g)
Mean S.D. (log10CFU/g)
Mean S.D. (log10CFU/g)
Infective endocarditis was induced in rats, using an inoculum of 56105 CFU containing GBS NCTC10/84 and its isogenic Dsrr1 mutant, at a 1:1 ratio. Animals were
sacrified 72 h post-infection, and log10 CFU/g of tissue for each strain was determined by plating onto selective media.
A number of bacterial surface structures have been shown to
mediate binding to fibrinogen, such as ClfA, ClfB, FnbA and Efb
of S. aureus, and the Fss proteins of Enterococcus faecalis . We
recently identified Srr1 of GBS as a fibrinogen-binding protein
that was important for bacterial attachment to microvascular
endothelial cells and CNS invasion . Although the binding
region of Srr1 has limited homology to other adhesins, analysis of
its predicted secondary structure indicated that the conformation
of this domain would resemble the binding region of ClfB. As has
been shown for several other adhesins, the binding pocket of ClfB
is formed by two Ig folds that engage the Aa chain of fibrinogen
via a dock, lock, and latch mechanism [35,39]. Srr1 appears to
interact with fibrinogen Aa by a similar mechanism, since deletion
of the predicted latch region abrogates fibrinogen binding by the
protein, and markedly reduces virulence in an animal model of
Our results indicate that Srr1-mediated binding to fibrinogen
also contributes to the pathogenesis of infective endocarditis.
Reduced fibrinogen binding in vitro was associated with decreased
virulence, as measured by our co-infection (competition) model of
endovascular infection. In particular, densities (CFU/g) of an Srr1
deletion mutant were significantly lower, as compared with its
parent strain, both within vegetations and in kidneys and spleens.
Of note, the mutant strain was not entirely avirulent, as it still
produced disease in the infected animals. This indicates that GBS
expresses other factors that contribute to its virulence, and is
consistent with other studies on the role of microbial binding in
endocarditis, where mutation or deletion of a single adhesin
produces only a partial reduction in pathogenicity [17,29,30].
GBS are known to express other fibrinogen binding proteins (FbsA
and FbsB), which may have contributed to the residual virulence
of our Dsrr-1 mutant strain [37,38]. Moreover it is likely that GBS
express additional surface components that can mediate binding to
cardiac valves, or enhance virulence by other mechanisms.
Binding to fibrinogen may be important for a number of events
in the pathogenesis of endovascular infections [10,40,41]. First,
bacterial attachment to the endocardium generally requires prior
alteration of the valve surface, such that it is covered with a matrix
of platelets and host proteins, including fibrinogen [20,4245].
Studies with S. aureus have shown that fibrinogen immobilized on
the valve surface is likely to contribute to the attachment of
circulating bacteria, thereby initiating infection . Our
current results indicate that fibrinogen may have a similar role for
GBS. In addition, fibrinogen in plasma could also serve to
crosslink GBS to platelets that have aggregated at sites of valve
injury. The subsequent progression of endovascular infection may
also be enhanced by GBS binding to fibrinogen. Bacteria
proliferating on the valve surface bacteria are thought to induce
the further deposition of fibrinogen onto the infected valve, which
in turn, is likely to trigger platelet attachment and aggregation.
These processes, in combination with bacterial growth, result in
the production of vegetations [9,10]. In view of our in vitro studies,
where Srr-1 enhanced the binding of bacteria to both fibrinogen
and platelets, is possible that Srr1-fibrinogen binding may be one
mechanism for the continued recruitment of platelets in vivo to the
infected endocardium, thereby stimulating disease progression.
A longstanding therapeutic goal has been to develop agents that
block bacterial binding to host tissues, thereby preventing or
attenuating subsequent infection. Fibrinogen binding is an
appealing target for disruption, in view of the importance of this
interaction for the pathogenesis of infective endocarditis. Although
inhibitory agents could target specific adhesins individually, such
as Srr1, an alternative strategy might be to develop drugs that
interfere with a larger number of dock, lock, and latch adhesins.
Although the binding clefts of these adhesins vary in terms of
primary amino acid sequence, it may still be possible to generate
agents that block binding, either by preventing docking, or by
inhibiting the latching process. If successful, this approach would
yield an inhibitor that could be used for a variety of pathogens.
We thank Dr. Barbara Bensing and Dr. Arnold S. Bayer for their helpful
scientific and editorial advice, and Dr. Kelly Doran for providing the GBS
isolates used in these studies.
Conceived and designed the experiments: HSS YQX PMS. Performed the
experiments: HSS YQX. Analyzed the data: HSS YQX PMS. Contributed
reagents/materials/analysis tools: HSS YQX PMS. Wrote the paper: HSS
1. Sunkara B , Bheemreddy S , Lorber B , Lephart PR , Hayakawa K , et al. ( 2012 ) Group B Streptococcus infections in non-pregnant adults: the role of immunosuppression . Int J Infect Dis 16 : e182 - 186 .
2. Farley MM ( 2001 ) Group B streptococcal disease in nonpregnant adults . Clin Infect Dis 33 : 556 - 561 .
3. Munoz P , Llancaqueo A , Rodriguez-Creixems M , Pelaez T , Martin L , et al. ( 1997 ) Group B streptococcus bacteremia in nonpregnant adults . Arch Intern Med 157 : 213 - 216 .
4. Siciliano RF , Cais DP , Navarro RC , Strabelli TM ( 2010 ) Acute Streptococcus agalactiae endocarditis: outcomes of early surgical treatment . Heart Lung 39 : 331 - 334 .
5. Ivanova Georgieva R , Garcia Lopez MV , Ruiz-Morales J , Martinez-Marcos FJ , Lomas JM , et al. ( 2010 ) Streptococcus agalactiae left-sided infective endocarditis. Analysis of 27 cases from a multicentric cohort . J Infect 61 : 54 - 59 .
6. Rollan MJ , San Roman JA , Vilacosta I , Sarria C , Lopez J , et al. ( 2003 ) Clinical profile of Streptococcus agalactiae native valve endocarditis . Am Heart J 146 : 1095 - 1098 .
7. Sambola A , Miro JM , Tornos MP , Almirante B , Moreno-Torrico A , et al. ( 2002 ) Streptococcus agalactiae infective endocarditis: analysis of 30 cases and review of the literature , 1962 - 1998 . Clin Infect Dis 34 : 1576 - 1584 .
8. Kannan R , Komaranchath AM , Mathew T , Ramprakash B , Sundararaman T , et al. ( 2001 ) Streptococcus agalactiae endocarditis . J Assoc Physicians India 49 : 1125 - 1126 .
9. Fitzgerald JR , Loughman A , Keane F , Brennan M , Knobel M , et al. ( 2006 ) Fibronectin-binding proteins of Staphylococcus aureus mediate activation of human platelets via fibrinogen and fibronectin bridges to integrin GPIIb/IIIa and IgG binding to the FcgammaRIIa receptor . Mol Microbiol 59 : 212 - 230 .
10. Loughman A , Fitzgerald JR , Brennan MP , Higgins J , Downer R , et al. ( 2005 ) Roles for fibrinogen, immunoglobulin and complement in platelet activation promoted by Staphylococcus aureus clumping factor A. Mol Microbiol 57 : 804 - 818 .
11. Fitzgerald JR , Foster TJ , Cox D ( 2006 ) The interaction of bacterial pathogens with platelets . Nat Rev Microbiol 4 : 445 - 457 .
12. Moreillon P , Que YA , Bayer AS ( 2002 ) Pathogenesis of streptococcal and staphylococcal endocarditis . Infect Dis Clin North Am 16 : 297 - 318 .
13. Veloso TR , Chaouch A , Roger T , Giddey M , Vouillamoz J , et al. ( 2012 ) Use of a Human-like Low-grade Bacteremia Model of Experimental Endocarditis to Study the Role of Staphylococcus aureus Adhesins and Platelet Aggregation in Early Endocarditis . Infect Immun 81 ( 3 ): 697 - 703 .
14. Zhou M , Wu H ( 2009 ) Glycosylation and biogenesis of a family of serine-rich bacterial adhesins . Microbiology 155 : 317 - 327 .
15. Bensing BA , Lopez JA , Sullam PM ( 2004 ) The Streptococcus gordonii surface proteins GspB and Hsa mediate binding to sialylated carbohydrate epitopes on the platelet membrane glycoprotein Ibalpha . Infect Immun 72 : 6528 - 6537 .
16. Lizcano A , Sanchez CJ , Orihuela CJ ( 2012 ) A role for glycosylated serine-rich repeat proteins in gram-positive bacterial pathogenesis . Mol Oral Microbiol 27 : 257 - 269 .
17. Siboo IR , Chambers HF , Sullam PM ( 2005 ) Role of SraP, a Serine-Rich Surface Protein of Staphylococcus aureus, in binding to human platelets . Infect Immun 73 : 2273 - 2280 .
18. Heilmann C , Niemann S , Sinha B , Herrmann M , Kehrel BE , et al. ( 2004 ) Staphylococcus aureus fibronectin-binding protein (FnBP)-mediated adherence to platelets, and aggregation of platelets induced by FnBPA but not by FnBPB . J Infect Dis 190 : 321 - 329 .
19. Miajlovic H , Loughman A , Brennan M , Cox D , Foster TJ ( 2007 ) Both complement- and fibrinogen-dependent mechanisms contribute to platelet aggregation mediated by Staphylococcus aureus clumping factor B . Infect Immun 75 : 3335 - 3343 .
20. Pietrocola G , Schubert A , Visai L , Torti M , Fitzgerald JR , et al. ( 2005 ) FbsA, a fibrinogen-binding protein from Streptococcus agalactiae, mediates platelet aggregation . Blood 105 : 1052 - 1059 .
21. Tazi A , Bellais S , Tardieux I , Dramsi S , Trieu-Cuot P , et al. ( 2012 ) Group B Streptococcus surface proteins as major determinants for meningeal tropism . Curr Opin Microbiol 15 : 44 - 49 .
22. Mistou MY , Dramsi S , Brega S , Poyart C , Trieu-Cuot P ( 2009 ) Molecular dissection of the secA2 locus of group B Streptococcus reveals that glycosylation of the Srr1 LPXTG protein is required for full virulence . J Bacteriol 191 : 4195 - 4206 .
23. van Sorge NM , Quach D , Gurney MA , Sullam PM , Nizet V , et al. ( 2009 ) The group B streptococcal serine-rich repeat 1 glycoprotein mediates penetration of the blood-brain barrier . J Infect Dis 199 : 1479 - 1487 .
24. Seifert KN , Adderson EE , Whiting AA , Bohnsack JF , Crowley PJ , et al. ( 2006 ) A unique serine-rich repeat protein (Srr-2) and novel surface antigen (epsilon) associated with a virulent lineage of serotype III Streptococcus agalactiae . Microbiology 152 : 1029 - 1040 .
25. Sheen TR , Jimenez A , Wang NY , Banerjee A , van Sorge NM , et al. ( 2011 ) Serine-rich repeat proteins and pili promote Streptococcus agalactiae colonization of the vaginal tract . J Bacteriol 193 : 6834 - 6842 .
26. Seo HS , Mu R , Kim BJ , Doran KS , Sullam PM ( 2012 ) Binding of glycoprotein Srr1 of Streptococcus agalactiae to fibrinogen promotes attachment to brain endothelium and the eevelopment of meningitis . PLoS Pathog 8 : e1002947 .
27. Seo HS , Xiong YQ , Mitchell J , Seepersaud R , Bayer AS , et al. ( 2010 ) Bacteriophage lysin mediates the binding of Streptococcus mitis to human platelets through interaction with fibrinogen . PLoS Pathog 6 : e1001047 .
28. Seo HS , Sullam PM ( 2011 ) Characterization of the fibrinogen binding domain of bacteriophage lysin from Streptococcus mitis . Infect Immun 79 : 3518 - 3526 .
29. Xiong YQ , Bensing BA , Bayer AS , Chambers HF , Sullam PM ( 2008 ) Role of the serine-rich surface glycoprotein GspB of Streptococcus gordonii in the pathogenesis of infective endocarditis . Microb Pathog 45 : 297 - 301 .
30. Mitchell J , Siboo IR , Takamatsu D , Chambers HF , Sullam PM ( 2007 ) Mechanism of cell surface expression of the Streptococcus mitis platelet binding proteins PblA and PblB . Mol Microbiol 64 : 844 - 857 .
31. Mitchell J , Sullam PM ( 2009 ) Streptococcus mitis phage-encoded adhesins mediate attachment to a2-8-linked sialic acid residues on platelet membrane gangliosides . Infect Immun 77 : 3485 - 3490 .
32. Que YA , Moreillon P ( 2011 ) Infective endocarditis . Nat Rev Cardiol 8 : 322 - 336 .
33. Geoghegan JA , Ganesh VK , Smeds E , Liang X , Hook M , et al. ( 2009 ) Molecular characterization of the interaction of staphylococcal microbial surface components recognizing adhesive matrix molecules (MSCRAMM) ClfA and Fbl with fibrinogen . J Biol Chem 285 : 6208 - 6216 .
34. Shannon O , Flock JI ( 2004 ) Extracellular fibrinogen binding protein, Efb, from Staphylococcus aureus binds to platelets and inhibits platelet aggregation . Thromb Haemost 91 : 779 - 789 .
35. Ganesh VK , Barbu EM , Deivanayagam CC , Le B , Anderson AS , et al. ( 2011 ) Structural and biochemical characterization of Staphylococcus aureus clumping factor B/ligand interactions . J Biol Chem 286 : 25963 - 25972 .
36. Sillanpaa J , Nallapareddy SR , Houston J , Ganesh VK , Bourgogne A , et al. ( 2009 ) A family of fibrinogen-binding MSCRAMMs from Enterococcus faecalis . Microbiology 155 : 2390 - 2400 .
37. Devi AS , Ponnuraj K ( 2010 ) Cloning, expression, purification and ligand binding studies of novel fibrinogen-binding protein FbsB of Streptococcus agalactiae . Protein Expr Purif 74 : 148 - 155 .
38. Pietrocola G , Visai L , Valtulina V , Vignati E , Rindi S , et al. ( 2006 ) Multiple interactions of FbsA, a surface protein from Streptococcus agalactiae, with fibrinogen: affinity, stoichiometry, and structural characterization . Biochemistry 45 : 12840 - 12852 .
39. Xiang H , Feng Y , Wang J , Liu B , Chen Y , et al. ( 2012 ) Crystal structures reveal the multi-ligand binding mechanism of Staphylococcus aureus ClfB . PLoS Pathog 8 : e1002751 .
40. O'Brien L , Kerrigan SW , Kaw G , Hogan M , Penades J , et al. ( 2002 ) Multiple mechanisms for the activation of human platelet aggregation by Staphylococcus aureus: roles for the clumping factors ClfA and ClfB, the serine-aspartate repeat protein SdrE and protein A. Mol Microbiol 44 : 1033 - 1044 .
41. Sullam PM , Valone FH , Mills J ( 1987 ) Mechanisms of platelet aggregation by viridans group streptococci . Infect Immun 55 : 1743 - 1750 .
42. Widmer E , Que YA , Entenza JM , Moreillon P ( 2006 ) New concepts in the pathophysiology of infective endocarditis . Curr Infect Dis Rep 8 : 271 - 279 .
43. Moreillon P , Que YA ( 2004 ) Infective endocarditis . Lancet 363 : 139 - 149 .
44. Nobbs AH , Lamont RJ , Jenkinson HF ( 2009 ) Streptococcus adherence and colonization . Microbiol Mol Biol Rev 73 : 407 - 450 .
45. Santoro J , Levison ME ( 1978 ) Rat model of experimental endocarditis . Infect Immun 19 : 915 - 918 .
46. Sullam PM , Bayer AS , Foss WM , Cheung AL ( 1996 ) Diminished platelet binding in vitro by Staphylococcus aureus is associated with reduced virulence in a rabbit model of infective endocarditis . Infect Immun 64 : 4915 - 4921 .
47. Xiong YQ , Fowler VG , Yeaman MR , Perdreau-Remington F , Kreiswirth BN , et al. ( 2009 ) Phenotypic and genotypic characteristics of persistent methicillinresistant Staphylococcus aureus bacteremia in vitro and in an experimental endocarditis model . J Infect Dis 199 : 201 - 208 .
48. Piroth L , Que YA , Widmer E , Panchaud A , Piu S , et al. ( 2008 ) The fibrinogenand fibronectin-binding domains of Staphylococcus aureus fibronectin-binding protein A synergistically promote endothelial invasion and experimental endocarditis . Infect Immun 76 : 3824 - 3831 .
49. Que YA , Haefliger JA , Piroth L , Francois P , Widmer E , et al. ( 2005 ) Fibrinogen and fibronectin binding cooperate for valve infection and invasion in Staphylococcus aureus experimental endocarditis . J Exp Med 201 : 1627 - 1635 .
50. Moreillon P , Entenza JM , Francioli P , McDevitt D , Foster TJ , et al. ( 1995 ) Role of Staphylococcus aureus coagulase and clumping factor in pathogenesis of experimental endocarditis . Infect Immun 63 : 4738 - 4743 .
51. Wessels MR , Haft RF , Heggen LM , Rubens CE ( 1992 ) Identification of a genetic locus essential for capsule sialylation in type III group B streptococci . Infect Immun 60 : 392 - 400 .
52. Wilkinson HW ( 1977 ) Nontypable group B streptococci isolated from human sources . J Clin Microbiol 6 : 183 - 184 .